Nicole Leibold

Prenatal stress and subsequent exposure to chronic mild stress in rats; interdependent effects on emotional behavior and the serotonergic system

Exposure to prenatal stress (PS) can predispose individuals to the development of psychopathology later in life. We examined the effects of unpredictable chronic mild stress (CMS) exposure during adolescence on a background of PS in male and female Sprague-Dawley rats. PS induced more anxiety-like behavior in the elevated zero maze in both sexes, an effect that was normalized by subsequent exposure to CMS. Moreover, PS was associated with increased depression-like behavior in the forced swim test in males only. Conversely, sucrose intake was increased in PS males, whilst being decreased in females when consecutively exposed to PS and CMS. Hypothalamo-pituitary-adrenal (HPA) axis reactivity was affected in males only, with higher stress-induced plasma corticosterone levels after PS. Markedly, CMS normalized the effects of PS on elevated zero maze behavior as well as basal and stress-induced plasma corticosterone secretion. At the neurochemical level, both PS and CMS induced various sex-specific alterations in serotonin (5-HT) and tryptophan hydroxylase 2 (TPH2) immunoreactivity in the dorsal raphe nucleus, hippocampus and prefrontal cortex with, in line with the behavioral observations, more profound effects in male offspring. In conclusion, these findings show that prenatal maternal stress in Sprague-Dawley rats induces various anxiety- and depression-related behavioral and neuroendocrine changes, as well as alterations in central 5-HT and TPH2 function, predominantly in male offspring. Moreover, CMS exposure partially normalized the effects of previous PS experience, suggesting that the outcome of developmental stress exposure largely depends on the environmental conditions later in life and vice versa.

Brainstem response to hypercapnia: A symptom provocation study into the pathophysiology of panic disorder

Background: The biological basis of uncued panic attacks is not yet understood. An important theory concerning the nature and cause of panic disorder is the ‘suffocation false alarm theory’. This alarm is supposed to be over-sensitive in panic disorder patients and can be triggered by CO2. No neurobiological substrate has been identified for such an alarm. The present study investigates differences in brain activation in panic patients, healthy individuals and experienced divers in response to CO2, representing three groups with descending sensitivity to CO2. Method: Brain activation was measured with functional magnetic resonance imaging. Subjects breathed through a mouthpiece delivering a continuous flow of 100% oxygen for two minutes, followed by a hypercapnic gas mixture (7% CO2) for the next two minutes. Statistical analysis was performed using SPM8. Results: There was a significant main effect of group in response to the CO2. Patients show increased brainstem activation in response to hypercapnia compared to controls and divers. Subjective feelings of breathing discomfort were positively correlated with brain activation in the anterior insula in all groups. Conclusion: This is the first study showing that the behavioural response to CO2 that characterises panic disorder patients is likely due to increased neural sensitivity to CO2 at brainstem level.

Carbon dioxide inhalation as a human experimental model of panic: the relationship between emotions and cardiovascular physiology.

Inhaling carbon dioxide (CO2)-enriched air induces fear and panic symptoms resembling real-life panic attacks, the hallmark of panic disorder. The present study aimed to describe the emotional and cardiovascular effects evoked by inhaling CO2, taking shortcomings of previous studies into account. Healthy volunteers underwent a double inhalation of 0, 9, 17.5, and 35% CO2, according to a randomized, cross-over design. In addition to fear, discomfort, and panic symptom ratings, blood pressure and heart rate were continuously monitored. Results showed a dose-dependent increase in all self-reports. Systolic and diastolic blood pressure rose with increasing CO2 concentration, whereas heart rate results were less consistent. Diastolic blood pressure and heart rate variation correlated with fear and discomfort. Based on this relationship and the observation that the diastolic blood pressure most accurately mimicked the degree of self-reported emotions, it might serve as a putative biomarker to assess the CO2-reactivity in the future.

Therapygenetics: 5-HTTLPR genotype predicts the response to exposure therapy for agoraphobia

This study was intended to assess the extent to which the low-expression allele of the serotonin transporter gene promoter predicts better response to exposure-based behavior therapy in patients with panic disorder with agoraphobia (PDA). Ninety-nine patients with PDA underwent a 1-week in vivo exposure-based behavior therapy program and provided saliva samples to extract genomic DNA and classify individuals according to four allelic forms (SA, SG, LA, LG) of the 5-HTT-linked polymorphic region (5-HTTLPR). We determined whether the 5-HTTLPR genotype predicted change in avoidance behavior in PDA following treatment. After controlling for pre-treatment avoidance behavior, the 5-HTTLPR low-expression genotypes showed a more favorable response to exposure therapy two weeks following treatment, compared to the other patients. This study suggests a genetic contribution to treatment outcome following behavior therapy and implicates the serotonergic system in response to exposure-based treatments in PDA.

CO2 exposure as translational cross-species experimental model for panic.

The current diagnostic criteria of the Diagnostic and Statistical Manual of Mental Disorders are being challenged by the heterogeneity and the symptom overlap of psychiatric disorders. Therefore, a framework toward a more etiology-based classification has been initiated by the US National Institute of Mental Health, the research domain criteria project. The basic neurobiology of human psychiatric disorders is often studied in rodent models. However, the differences in outcome measurements hamper the translation of knowledge. Here, we aimed to present a translational panic model by using the same stimulus and by quantitatively comparing the same outcome measurements in rodents, healthy human subjects and panic disorder patients within one large project. We measured the behavioral–emotional and bodily response to CO2 exposure in all three samples, allowing for a reliable cross-species comparison. We show that CO2 exposure causes a robust fear response in terms of behavior in mice and panic symptom ratings in healthy volunteers and panic disorder patients. To improve comparability, we next assessed the respiratory and cardiovascular response to CO2, demonstrating corresponding respiratory and cardiovascular effects across both species. This project bridges the gap between basic and human research to improve the translation of knowledge between these disciplines. This will allow significant progress in unraveling the etiological basis of panic disorder and will be highly beneficial for refining the diagnostic categories as well as treatment strategies.

The brain acid-base homeostasis and serotonin: A perspective on the use of carbon dioxide as human and rodent experimental model of panic.

Panic attacks (PAs), the core feature of panic disorder, represent a common phenomenon in the general adult population and are associated with a considerable decrease in quality of life and high health care costs. To date, the underlying pathophysiology of PAs is not well understood. A unique feature of PAs is that they represent a rare example of a psychopathological phenomenon that can be reliably modeled in the laboratory in panic disorder patients and healthy volunteers. The most effective techniques to experimentally trigger PAs are those that acutely disturb the acid-base homeostasis in the brain: inhalation of carbon dioxide (CO2), hyperventilation, and lactate infusion. This review particularly focuses on the use of CO2 inhalation in humans and rodents as an experimental model of panic. Besides highlighting the different methodological approaches, the cardio-respiratory and the endocrine responses to CO2 inhalation are summarized. In addition, the relationships between CO2 level, changes in brain pH, the serotonergic system, and adaptive physiological and behavioral responses to CO2 exposure are presented. We aim to present an integrated psychological and neurobiological perspective. Remaining gaps in the literature and future perspectives are discussed.

The anatomy of fear learning in the cerebellum: A systematic meta-analysis.

Recent neuro-imaging studies have implicated the cerebellum in several higher-order functions. Its role in human fear conditioning has, however, received limited attention. The current meta-analysis examines the loci of cerebellar contributions to fear conditioning in healthy subjects, thus mapping, for the first time, the neural response to conditioned aversive stimuli onto the cerebellum. By using the activation likelihood estimation (ALE) technique for analyses, we identified several distinct regions in the cerebellum that activate in response to the presentation of the conditioned stimulus: the cerebellar tonsils, lobules HIV-VI, and the culmen. These regions have separately been implicated in fear acquisition, consolidation of fear memories and expression of conditioned fear responses. Their specific role in these processes may be attributed to the general contribution of cerebellar cortical networks to timing and prediction. Our meta-analysis highlights the potential role of the cerebellum in human cognition and emotion in general, and addresses the possibility how deficits in associative cerebellar learning may play a role in the pathogenesis of anxiety disorders. Future studies are needed to further clarify the mechanistic role of the cerebellum in higher order functions and neuropsychiatric disorders.

Amiloride-sensitive cation channel 2 genotype affects the response to a carbon dioxide panic challenge

Until recently, genetic research into panic disorder (PD) has had only limited success. Inspired by rodent research, demonstrating that the acid-sensing ion channel 1a (ASIC1a) is critically involved in the behavioral fear response to carbon dioxide (CO2) exposure, variants in the human homologue gene amiloride-sensitive cation channel 2 (ACCN2) were shown to be associated with PD. However, the relationship between changes in brain pH and ACCN2, as done in rodents by CO2 exposure, has not been investigated yet in humans. Here, we examined this link between the ACCN2 gene and the response to CO2 exposure in two studies: in healthy volunteers as well as PD patients and using both behavioral and physiological outcome measures. More specifically, 107 healthy volunteers and 183 PD patients underwent a 35% CO2 inhalation. Negative affect was assessed using visual analogue scales and the panic symptom list (PSL), and, in healthy volunteers, cardiovascular measurements. The single nucleotide polymorphism rs10875995 was significantly associated with a higher emotional response in PD patients and with an increase in systolic as well as diastolic blood pressure in healthy subjects. In all measurements, subjects homozygous for the T-allele showed a heightened reactivity to CO2. Furthermore, a trend towards an rs685012 genotype effect on the emotional response was found in PD patients. We provide the first evidence that genetic variants in the ACCN2 are associated with differential sensitivity to CO2 in PD patients as well as healthy volunteers, further supporting ACCN2 as a promising candidate for future research to improve current treatment options.

Behavioral pattern separation and its link to the neural mechanisms of fear generalization

Abstract Fear generalization is a prominent feature of anxiety disorders and post-traumatic stress disorder (PTSD). It is defined as enhanced fear responding to a stimulus that bears similarities, but is not identical to a threatening stimulus. Pattern separation, a hippocampal-dependent process, is critical for stimulus discrimination; it transforms similar experiences or events into non-overlapping representations. This study is the first in humans to investigate the extent to which fear generalization relies on behavioral pattern separation abilities. Participants (N = 46) completed a behavioral task taxing pattern separation, and a neuroimaging fear conditioning and generalization paradigm. Results show an association between lower behavioral pattern separation performance and increased generalization in shock expectancy scores, but not in fear ratings. Furthermore, lower behavioral pattern separation was associated with diminished recruitment of the subcallosal cortex during presentation of generalization stimuli. This region showed functional connectivity with the orbitofrontal cortex and ventromedial prefrontal cortex. Together, the data provide novel experimental evidence that pattern separation is related to generalization of threat expectancies, and reduced fear inhibition processes in frontal regions. Deficient pattern separation may be critical in overgeneralization and therefore may contribute to the pathophysiology of anxiety disorders and PTSD.

Brain and Behavior Changes following Exposure Therapy Predict Outcome at 8-Year Follow-Up.

local ethics committee approved the study, and written consent was obtained. Behavioral change was examined with repeated measures analyses of variance. Short-term behavioral changes (T1–T0) were correlated with long-term outcome (SPQ). For the fMRI analyses in SPM8, we first created individual contrast images for spider > neutral, followed by statistical images of therapy-related change (T1– T0). A second-level regression analysis was used to evaluate baseline and changes (T1–T0) in neural activation associated with follow-up SPQ scores. A familywise error rate small-volume correction was applied for a priori regions of interest: left amygdala, anterior cingulate cortex, and left insula, as these regions showed therapy-induced changes [2] . Additionally, an exploratory whole-brain second-level analysis was conducted. Figure 1 shows an overview of the behavioral and neural results. Although SPQ scores at follow-up were increased compared to SPQ scores at T1 (p < 0.01), they were still lower than T0 ratings [F(1.37, 19.19) = 57.342; p < 0.001; post hoc p < 0.001], indicating that the therapy effect was partially maintained over 8 years. Furthermore, SPQ and fear/arousal score reductions from preto posttherapy assessments (T1–T0; r = 0.78, p < 0.01; r = 0.57, p < 0.05; r = 0.70, p < 0.01) were associated with a lower SPQ score at follow-up. Similarly, a lower SPQ score at follow-up was associated with more attenuation in the BOLD response from preto posttherapy time points (T1–T0) in the left anterior insula (cluster extending into the lateral orbitofrontal cortex, peak voxel MNI coordinates –26/24/–12, cluster size = 29 voxels, t = 7.20, p < 0.01, r = 0.89). This effect was not driven by changes in processing of neutral images between T0 and T1. Change in activation (T1–T0) in other brain regions did not correlate with the follow-up SPQ scores, as tested by the small-volume correction analysis and whole-brain analysis. To check whether the observed associations were driven by T1–T0 variance, we further tested whether associations existed between the follow-up SPQ score and behavioral and neural data at T0 and T1. We only found a correlation with the T1 SPQ score and the follow-up SPQ score (r = 0.91, p < 0.001). In conclusion, the present findings show strong negative correlations between phobia severity at the 8-year follow-up and changes (T1–T0) in anterior insula activation and fear scores. Recently, it has been argued that symptom reduction directly following exposure therapy for phobia is only weakly predictive of treatment outcome [3] . The present findings, however, point to the usefulness of assessing fear levels in the evaluation of the therapy effect when not measured immediately after therapy, but after allowing time for consolidation of learning. The demonstrated association between the extent of insular activity reduction and better clinical outcome at 8 years of followEven the most effective psychological treatments, such as cognitive behavioral therapy, show high percentages of relapse in the long term [1] . Predicting the persistence of therapy effects remains a major unanswered question, which is complicated by psychiatric comorbidity and by limited insight into the effectiveness of different therapy components applied during cognitive behavioral therapy. Neuroimaging offers the possibility of objective measures for predicting long-term therapy success and to elucidate its mechanisms. A first step to disentangle the problem of predicting therapy persistence is to (1) investigate a phenotypically clear disorder like specific phobia and (2) choose one cognitive behavioral therapy component targeting a single therapeutic mechanism such as exposure therapy, the clinical application of extinction learning [1] . Here, we link behavioral and neural changes following exposure therapy for specific phobia with behavioral measures at 8-year follow-up, to investigate which outcome measures of short-term therapeutic assessment are associated with long-term persistence of therapeutic response. Sixteen right-handed, unmedicated female spider phobic subjects received one prolonged (3–5 h) session of group-based exposure therapy. The diagnosis of specific phobia was made by an experienced psychiatrist (K.S.) based on DSM-IV criteria. Any further psychopathology was excluded by means of a structured psychiatric interview. Subjects were scanned twice, 1 week before (T0) and 2 weeks after therapy (T1), while undergoing a symptom provocation functional magnetic resonance imaging (fMRI) paradigm. Behavioral ratings at T0 and T1 consisted of phobia severity (Spider Phobia Questionnaire, SPQ) and fear/arousal for spiders [2] . At 8 years of follow-up, 15 subjects agreed to participate in the follow-up study and rated phobia severity on the SPQ again. The Received: July 23, 2015 Accepted after revision: November 6, 2015 Published online: May 27, 2016